MPK4 Phosphorylation Dynamics and Interacting Proteins in Plant Immunity

Arabidopsis MAP
kinase 4 (MPK4) has been proposed
to be a negative player in plant immunity, and it is also activated
by pathogen-associated molecular patterns (PAMPs), such as flg22.
The molecular mechanisms by which MPK4 is activated and regulates
plant defense remain elusive. In this study, we investigated Arabidopsis defense against a bacterial pathogen Pseudomonas syringae pv tomato (Pst) DC3000
when Brassica napus MPK4 (BnMPK4) is overexpressed. We showed an increase in pathogen resistance
and suppression of jasmonic acid (JA) signaling in the BnMPK4 overexpressing (OE) plants. We also showed that the OE plants have
increased sensitivity to flg22-triggered reactive oxygen species (ROS)
burst in guard cells, which resulted in enhanced stomatal closure
compared to wild-type (WT). During flg22 activation, dynamic phosphorylation
events within and outside of the conserved TEY activation loop were
observed. To elucidate how BnMPK4 functions during
the defense response, we used immunoprecipitation coupled with mass
spectrometry (IP-MS) to identify BnMPK4 interacting
proteins in the absence and presence of flg22. Quantitative proteomic
analysis revealed a shift in the MPK4-associated protein network,
providing insight into the molecular functions of MPK4 at the systems
level.